The objectives of this U01 project are to work with the Countermeasures Against Chemical Threats (CounterACT) Research Network to establish proof of concept that PEGylated recombinant human butyrylcholinesterase (Protexia), a bioscavenger that neutralizes organophosphate nerve agents in vivo, is efficacious as a therapy following lethal exposure to nerve agent. Protexia will be delivered intramuscularly in a bioactive form with a small preloaded autoinjector that is suitable for rapid first responder administration in a mass casualty setting. The Specific Aims of this project are: 1) To demonstrate therapeutic efficacy of Protexia in guinea pigs after exposure to VX. All animal studies with VX will be conducted at Defense Research and Development Canada (DRDC Suffield, Canada), which is certified to test chemical agent mitigation systems in rodent and large animal models. 2) To demonstrate therapeutic efficacy of IV Protexia in anesthetized domestic pigs after exposure to VX. 3) To demonstrate rapid Protexia delivery into the systemic circulation with a commercially available intramuscular autoinjector from Meridian Medical Technologies, which produces nerve agent antidote autoinjectors for military and civilian use. 4) To optimize and improve the autoinjector IM delivery of bioactive Protexia. 5) To demonstrate that protective serum levels of Protexia can be achieved in domestic pigs by autoinjector delivery. 6) To demonstrate that autoinjected Protexia can be used to treat domestic pigs following a lethal challenge with VX. The primary endpoint will be survival and the secondary endpoint will be the inhibition of neurological sequelae. 7) To perform a safety study of Protexia in non-human primates that will enable the submission of an Investigational New Drug (IND) application to the FDA. This study will be carried out in cynomolgus monkeys by Charles River Laboratories, Inc (Sparks, NV). Relevance. The overall goal of this research program is to develop a novel therapeutic agent to treat large numbers of persons exposed to nerve agents as a result of a mass civilian attack. Pharm- Athene has produced recombinant human butyrylcholinesterase (rBChE) at commercially feasible levels in the milk of transgenic goats and demonstrated that pre-exposure prophylactic treatment with PEGylated rBChE (Protexia) protects animals against lethal doses of nerve agents such as soman and VX. Preliminary data suggests that Protexia may be protective if administered after mass exposure to nerve agent. [unreadable] [unreadable] [unreadable]